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J Biol Chem. 2018 Mar 16;293(11):4026-4036. doi: 10.1074/jbc.RA117.001642. Epub 2018 Jan 30.

Structure-function analyses of a PL24 family ulvan lyase reveal key features and suggest its catalytic mechanism.

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From the Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E5, Canada.
the Université Grenoble Alpes and CNRS, CERMAV UPR 5301 601, rue de la chimie, 38000 Grenoble (France) and Institut de Chimie Moléculaire de Grenoble, ICMG, FR-CNRS 2607, Grenoble, France.
the Institute for Nanotechnology and Advanced Materials, and Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52900, Israel, and.
From the Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5E5, Canada,
the Department of Biochemistry, McGill University, Montreal, Quebec H3G 0B1, Canada.


Ulvan is a major cell wall component of green algae of the genus Ulva, and some marine bacteria encode enzymes that can degrade this polysaccharide. The first ulvan-degrading lyases have been recently characterized, and several putative ulvan lyases have been recombinantly expressed, confirmed as ulvan lyases, and partially characterized. Two families of ulvan-degrading lyases, PL24 and PL25, have recently been established. The PL24 lyase LOR_107 from the bacterial Alteromonadales sp. strain LOR degrades ulvan endolytically, cleaving the bond at the C4 of a glucuronic acid. However, the mechanism and LOR_107 structural features involved are unknown. We present here the crystal structure of LOR_107, representing the first PL24 family structure. We found that LOR_107 adopts a seven-bladed β-propeller fold with a deep canyon on one side of the protein. Comparative sequence analysis revealed a cluster of conserved residues within this canyon, and site-directed mutagenesis disclosed several residues essential for catalysis. We also found that LOR_107 uses the His/Tyr catalytic mechanism, common to several PL families. We captured a tetrasaccharide substrate in the structures of two inactive mutants, which indicated a two-step binding event, with the first substrate interaction near the top of the canyon coordinated by Arg320, followed by sliding of the substrate into the canyon toward the active-site residues. Surprisingly, the LOR_107 structure was very similar to that of the PL25 family PLSV_3936, despite only ∼14% sequence identity between the two enzymes. On the basis of our structural and mutational analyses, we propose a catalytic mechanism for LOR_107 that differs from the typical His/Tyr mechanism.


His/Tyr mechansim; Polysaccharide lyase; Ulvan lyase; carbohydrate processing; catalysis; crystallography; mutagenesis; polysaccharide; structural biology; substrate specificity

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